Abstract How human cardiomyocytes (hCMs) acquire and maintain cardiac fate remains largely unknown due to the limited availability of genetic, molecular and cellular tools. Increasing evidence, however, indicates significant differences between human and mouse CM fate acquisition, suggesting the limitation of using only animal model to study human heart development and diseases. Programming pluripotent stem cells and reprogramming somatic cells such as fibroblasts into cardiac fate provide valuable platforms for studying hCM cell fate determination, disease modeling, drug screen and in vivo repair. Over the years, we have established a robust murine induced cardiomyocyte (iCM) reprogramming system. By using this system, we obtained novel insights into the transcriptional, posttranscriptional and epigenetic regulation of murine CM fate, and concomitantly improved the quality and yield of murine iCMs. In this research program, we will exploit our expertise in iCM reprogramming and single cell omics as well as our newly acquired preliminary data to define the molecular routes of, and epigenetic and metabolic mechanisms underlying human iCM conversion. Successful completion of this proposal will help to move direct cardiac reprogramming closer to its clinical application, and provide new insights into molecular mechanisms underlying human cardiomyocyte fate acquisition and maintenance.